Project 1. Clinical Trials Abstract Non-small cell lung cancer (NSCLC) and Malignant Pleural mesothelioma (MPM) are highly lethal cancers with suboptimal treatment. The treatment strategy to be used in this project is to produce genetically engineered T cells expressing chimeric antigen receptors (CARs) that target T cells to tumors. CAR T cells have shown impressive early clinical results in hematologic malignancies. We were among the first groups in the world to inititate CAR T cell trials focused on solid tumors by targeting the tumor-associated antigen mesothelin. Despite showing safety and feasibility, efficacy was limited, a common theme in solid tumors. In this Project, our goal is to use information from our previous clinical trials and ongoing studies in Projects 2 and 3 to conduct new CAR T cell clinical trials in patients with lung cancer and MPM. Our recent data suggest that two key problems in our previous trials were suboptimal anti-tumor activity and a rapid immune-based rejection of the CAR T cells due to the murine origin of the single chain antibody on the CAR. Accordingly, in Aim 1, we will complete a clinical trial in patients with NSCLC and MPM using a newly designed anti-mesothelin-CAR construct, called huCART-meso, that contains a fully human single chain antibody and has much higher activity in preclinical models than the previously used SS1 CAR. We will also evaluate the possible benefits of: 1) pre-treatment lymphodepletion using cyclophosphamide and 2) local delivery of CAR T cells in patients with malignant pleural effusions. Other barriers to solid tumor CAR therapy success are likely to be tumor heterogeneity, whereby not all tumor cells express the targeted antigen, poor trafficking of T cells, and local immunosuppression. An approach to these issues is to attack the tumor stroma. Previous studies from our group and others support the hypothesis that CAR T cells targeting a protein called Fibroblast Activation Protein (FAP) can lower the number of cancer- associated fibroblasts, reduce tumor size and stimulate anti-tumor immunity. In Aim 2, we will test this approach in patients by conducting a Phase 1 clinical trial to evaluate the potential safety and clinical activity of a CAR targeted to human FAP. In both Aims, we will also address the issue of tumor heterogeneity by providing samples to Project 3 where we will study the ability of CAR T cells to induce epitope spreading, a process by which the CAR T cells kill tumor cells in an immunogenic fashion and induce endogenous anti- tumor T cell or B cell activity. Based on the results of these two trials and on new data from Projects 2 and 3, in future studies, we will design a third CAR T cell trial. Trials we will consider include: 1) adding checkpoint blockade, 2) combining huCART-meso and FAPCAR T cells, and 3) using even more highly active CARs. The PO1 and Project 1 have high significance: achieving success rates with CAR T cells in solid tumors similar to that observed in leukemia would be a paradigm shift in the treatment of solid tumors. 1